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Patatin-like phospholipase domain-containing lipase 8 (PNPLA8), one of the calcium-independent phospholipase A2 enzymes, is involved in various physiological processes through the maintenance of membrane phospholipids. Biallelic variants in PNPLA8 have been associated with a range of paediatric neurodegenerative disorders. However, the phenotypic spectrum, genotype-phenotype correlations and the underlying mechanisms are poorly understood. Here, we newly identified 14 individuals from 12 unrelated families with biallelic ultra-rare variants in PNPLA8 presenting with a wide phenotypic spectrum of clinical features. Analysis of the clinical features of current and previously reported individuals (25 affected individuals across 20 families) showed that PNPLA8-related neurological diseases manifest as a continuum ranging from variable developmental and/or degenerative epileptic-dyskinetic encephalopathy to childhood-onset neurodegeneration. We found that complete loss of PNPLA8 was associated with the more profound end of the spectrum, with congenital microcephaly. Using cerebral organoids generated from human induced pluripotent stem cells, we found that loss of PNPLA8 led to developmental defects by reducing the number of basal radial glial cells and upper-layer neurons. Spatial transcriptomics revealed that loss of PNPLA8 altered the fate specification of apical radial glial cells, as reflected by the enrichment of gene sets related to the cell cycle, basal radial glial cells and neural differentiation. Neural progenitor cells lacking PNPLA8 showed a reduced amount of lysophosphatidic acid, lysophosphatidylethanolamine and phosphatidic acid. The reduced number of basal radial glial cells in patient-derived cerebral organoids was rescued, in part, by the addition of lysophosphatidic acid. Our data suggest that PNPLA8 is crucial to meet phospholipid synthetic needs and to produce abundant basal radial glial cells in human brain development., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Guarantors of Brain.)

Angelman syndrome (AS) is a severe neurodevelopmental disorder caused by the loss of function of maternal UBE3A. The major cause of AS is a maternal deletion in 15q11.2-q13, and the minor causes are a UBE3A mutation, uniparental disomy (UPD), and imprinting defect (ID). Previous reports suggest that all patients with AS exhibit developmental delay, movement or balance disorders, behavioral characteristics, and speech impairment. In contrast, a substantial number of AS patients with a UBE3A mutation, UPD, or ID were reported not to show these consistent features and to show age-dependent changes in their features. In this study, we investigated 134 patients with AS, including 57 patients with a UBE3A mutation and 48 patients with UPD or ID. Although developmental delay was present in all patients, 20% of patients with AS caused by UPD or ID did not exhibit movement or balance disorders. Differences were also seen in hypopigmentation and seizures, depending on the causes. Moreover, patients with a UBE3A mutation, UPD, or ID tended to show fewer of the specific phenotypes depending on their age. In particular, in patients with UPD or ID, easily provoked laughter and hyperactivity tended to become more pronounced as they aged. Therefore, the clinical features of AS based on cause and age should be understood, and genetic testing should not be limited to patients with the typical clinical features of AS., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Objectives: The 18- and 16-mm conduits in extracardiac total cavopulmonary connection (eTCPC) were reported to be optimal based on energy loss and flow stagnation at the relatively early phase. However, because the artificial conduit lacks growth potential, we have recently encountered some cases in which the conduit needs to be changed several years after eTCPC. These cases prompted us to reconsider the surgical strategy for eTCPC., Methods: We reviewed our 20-year single-centre experience with eTCPC patients (n = 256) to compare the 18-mm conduit (n = 195) and 16-mm conduit (n = 61) in terms of mortality and morbidity., Results: The 16-mm conduit was used significantly more frequently in patients whose main chamber was right ventricle (P < 0.001). There was also a significant difference in preoperative inferior vena cava pressure (P = 0.008). There was a significant difference in the actuarial rate of freedom from late-occurring complications, including mortality, between the 2 groups (P = 0.003). There was a significant difference in the actuarial rate of reoperation-free survival (P = 0.042); however, there was no significant difference in resurgical intervention for the conduit (P = 0.333). In multivariate analysis, preoperative inferior vena cava pressure was an independent predictor for late-occurring complications (hazard ratio 1.19; P = 0.026). Conduit size (18 or 16 mm) itself was not an independent predictive factor for late-occurring complications (P = 0.690)., Conclusions: The mid-term clinical outcomes in patients who underwent eTCPC were excellent with low mortality. Preoperative inferior vena cava pressure was the only predictive risk factor for postoperative morbidity, and the 16 mm conduit was not predictive thereof., (© The Author(s) 2024. Published by Oxford University Press on behalf of the European Association for Cardio-Thoracic Surgery.)

Reoperation after pediatric mitral valve replacement (MVR) is inevitable due to patient-prosthesis mismatch (PPM) associated with somatic growth. We analyzed potential metrics for PPM and outcomes of redo MVR for valve upsizing. Between 1999 and 2018, 15 children without obstructive left heart lesions other than mitral stenosis underwent initial MVR with a 16-mm ATS-Advanced Performance valve. We analyzed hemodynamic data from 28 postoperative catheterizations and concomitant echocardiograms. The median age and body weight at initial MVR were 4.9 months (25th, 75th percentile: 3.6, 6.6) and 5.9 kg (5.0, 7.3). Redo MVR was planned when patients had congestive heart failure and postcapillary pulmonary hypertension (PH) due to PPM: systolic pulmonary arterial pressure (SPAP) >35 mm Hg and pulmonary capillary wedge pressure (PCWP) >15 mm Hg on catheterization. Indexed effective orifice area (iEOA) and mean transmitral pressure gradient (TMPG) were strongly correlated with SPAP (r = -0.72, P < 0.001 and r = 0.75, P < 0.001) and PCWP (r = -082, P < 0.001 and r = 0.84, P < 0.001). Cut-off values for detecting postcapillary PH due to PPM were 1.0 cm 2 /m 2 for iEOA and 18 mm Hg for mean TMPG. Nine patients underwent redo MVR for postcapillary PH due to PPM at a median postoperative interval of 10 years (9.2, 11.9). All the patients survived, and PH was improved one year after surgery. iEOA and mean TMPG can be metrics for PPM in children after MVR. Careful follow-up is required to confirm the improvement of preoperatively existing PH after redo MVR for valve upsizing., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Background: In Kampo medicine, tongue examination is used to diagnose the pathological condition "Sho," but an objective evaluation method for its diagnostic ability has not been established. We constructed a tongue diagnosis electronic learning and evaluation system based on a standardized tongue image database., Purpose: This study aims to verify the practicality of this assessment system by evaluating the tongue diagnosis ability of Kampo specialists (KSs), medical professionals, and students., Methods: In the first study, we analyzed the answer data of 15 KSs in an 80-question tongue diagnosis test that assesses eight aspects of tongue findings and evaluated the (i) test score, (ii) test difficulty and discrimination index, (iii) diagnostic consistency, and (iv) diagnostic match rate between KSs. In the second study, we administered a 20-question common Kampo test and analyzed the answer data of 107 medical professionals and 56 students that assessed the tongue color discrimination ability and evaluated the (v) correct answer rate, (vi) test difficulty, and (vii) factors related to the correct answer rate., Result: In the first study, the average test score was 62.2 ± 10.7 points. Twenty-eight questions were difficult (correct answer rate, <50%), 34 were moderate (50%-85%), and 18 were easy (≥85%). Regarding intrarater reliability, the average diagnostic match rate of five KSs involved in database construction was 0.66 ± 0.08, and as for interrater reliability, the diagnostic match rate between the 15 KSs was 0.52 (95% confidence interval, 0.38-0.65) for Gwet's agreement coefficient 1, and the degree of the match rate was moderate. In the second study, the difficulty level of questions was moderate, with a correct rate of 81.3% for medical professionals and 82.1% for students. The discrimination index was good for medical professionals (0.35) and poor for students (0.06). Among medical professionals, the correct answer group of this question had a significantly higher total score on the Kampo common test than the incorrect answer group (85.3 ± 8.4 points vs. 75.8 ± 11.8 points, p  < 0.01)., Conclusion: This system can objectively evaluate tongue diagnosis ability and has high practicality. Utilizing this system can be expected to contribute to improving learners' tongue diagnosis ability and standardization of tongue diagnosis., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (© 2023 Segawa, Iizuka, Ogihara, Tanaka, Nakae, Usuku, Yamaguchi, Wada, Uchizono, Nakamura, Nishida, Ueda, Shiota, Hasunuma, Nakahara, Hebiguchi and Hamamoto.)

Angelman syndrome (AS) is caused by the functional absence of the maternal ubiquitin-protein ligase E3A (UBE3A) gene. Approximately 5% of AS is caused by paternal uniparental disomy of chromosome 15 (UPD(15)pat), most of which is considered to result from monosomy rescue. However, little attention has focused on how UPD(15)pat occurs. We suggest the mitotic nondisjunction mechanism as a cause of UPD(15)pat in a six-year-old patient presenting with distinctive characteristics in line with AS. DNA methylation screening of 15q11-q13 showed a paternal band and a faint maternal band, suggestive of mosaic status. By trio-based microsatellite analysis, we confirmed a large proportion of UPD(15)pat cells and a small proportion of cells of biparental origin. Single nucleotide polymorphism (SNP) microarray revealed isodisomy of the entire chromosome 15. These results suggest that the UPD(15)pat of the patient resulted from mitotic nondisjunction, which may also be the cause of other cases of AS with UPD(15)pat., (© 2022. The Author(s), under exclusive licence to The Japan Society of Human Genetics.)

For plasma spectroscopy, Stokes spectropolarimetry is used as a method to spatially invert the viewing-chord-integrated spectrum on the basis of the correspondence between the given magnetic field profile along the viewing chord and the Zeeman effect appearing on the spectrum. Its application to fusion-related toroidal plasmas is, however, limited owing to the low spatial resolution as a result of the difficulty in distinguishing between the Zeeman and Doppler effects. To resolve this issue, we increased the relative magnitude of the Zeeman effect by observing a near-infrared emission line on the basis of the greater wavelength dependence of the Zeeman effect than of the Doppler effect. By utilizing the increased Zeeman effect, we are able to invert the measured spectrum with a high spatial resolution by Monte Carlo particle transport simulation and by reproducing the measured spectra with the semiempirical adjustment of the recycling condition at the first walls. The inversion result revealed that when the momentum exchange collisions of atoms are negligible, the velocity distribution of core-fueling atoms is mainly determined by the initial distribution at the time of recycling. The inversion result was compared with that obtained using a two-point emission model used in previous studies. The latter approximately reflects the parameters of atoms near the emissivity peak., (© 2022. The Author(s).)

Recently, whole-exome sequencing (WES) has been used for genetic diagnoses of patients who remain otherwise undiagnosed. WES was performed in 177 Japanese patients with undiagnosed conditions who were referred to the Tokai regional branch of the Initiative on Rare and Undiagnosed Diseases (IRUD) (TOKAI-IRUD). This study included only patients who had not previously received genome-wide testing. Review meetings with specialists in various medical fields were held to evaluate the genetic diagnosis in each case, which was based on the guidelines of the American College of Medical Genetics and Genomics. WES identified diagnostic single-nucleotide variants in 66 patients and copy number variants (CNVs) in 11 patients. Additionally, a patient was diagnosed with Angelman syndrome with a complex clinical phenotype upon detection of a paternally derived uniparental disomy (UPD) [upd(15)pat] wherein the patient carried a homozygous DUOX2 p.E520D variant in the UPD region. Functional analysis confirmed that this DUOX2 variant was a loss-of-function missense substitution and the primary cause of congenital hypothyroidism. A significantly higher proportion of genetic diagnoses was achieved compared to previous reports (44%, 78/177 vs. 24-35%, respectively), probably due to detailed discussions and the higher rate of CNV detection., (© 2022. The Author(s).)

Alpha-thalassemia X-linked intellectual disability (ATR-X) syndrome is caused by a mutation in ATRX, which is essential for proper chromatin remodeling. ATRX dysfunction leads to dysregulation of many genes due to abnormal chromatin remodeling, and causes a multisystem disorder in patients with ATR-X. Because mitochondrial disorders also show multisystem involvement, whether mitochondrial function is affected in patients with ATR-X is of interest. Here, we report a case of a 4-year-old male with a mutation (NM&#95;000489.4: c.736C > T p.Arg246Cys) in ATRX, who showed mitochondrial dysfunction with complex I deficiency. The results from our study suggest that target genes of the ATRX protein may include those responsible for mitochondrial function, and mitochondrial dysfunction may contribute to some ATR-X phenotypes., (Copyright © 2021 Elsevier Masson SAS. All rights reserved.)

Acoustic wave can destabilize the flame and has a potential in firefighting, but the influences of the sound source and its frequency are still poorly understood. This work applies a loudspeaker to extinguish a laminar diffusion propane flame of 5-25 mm high, where the local sound frequency is 50-70 Hz and sound pressure is 0.8-3.2 Pa (92.0-104.1 dB). Results reveal a constant flame pulsating displacement at the extinction limit, independent of the sound environment used. Such a flame pulsating displacement is found to be caused by the motion of the speaker membrane (or diaphragm) and its induced wind, which could be two orders of magnitude larger than the displacement of the air that transmits acoustic wave. Thus, under the influence of sound source, a critical flame strain rate, stretched by the pulsating airflow, can be formulated to characterize the blow-off limit better than the local sound pressure. The sound source with a lower frequency can produce larger pulsating displacements of both membrane and flame, and thus promoting extinction. This work improves the understanding of flame dynamics under the external sound field and source, and it helps establish a scientific framework for acoustic-based fire suppression technologies., (© 2021. The Author(s).)

The elution of lead, and nickel from water supply devices into water is a potential health concern. This study was performed to examine the actual concentrations of nickel and lead in the water from taps in homes and offices, focusing on the differences between first flush and fully flushed water. The water quality management target value and water quality standard in Japan specify nickel and lead concentrations in drinking water <20 and <10 μg/L, respectively. Nickel concentration in the first flush water (100 mL) from 110 household taps revealed 22 cases (20%) > 20 μg/L, while the fully flushed water satisfied the standard after running 5000 mL of water. The nickel concentration decreased gradually in sequential sampling of each 100 mL from the taps. Lead concentration in the first flush water exceeded the standard in 32 cases (29%), while the fully flushed water was below the target value. The concentration in the first flush water tended to decrease with time since the tap installation, and this was significant after 10 years for nickel but not significant for lead. It is important to flush retained water out of the tap after several hours without use. No significant correlation was found with the volume of the test faucet in the market, but bronze-based products showed higher nickel concentrations than brass and plastic products., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021. Published by Elsevier B.V.)

A 1-month-old girl with common arterial trunk, anteriorly originated crisscross pulmonary arteries, unusual running right aortic arch and severe pulmonary hypertension was initially palliated by bilateral pulmonary artery banding. She developed sudden respiratory failure at 80 days old; and computed tomography revealed that the right main bronchus was severely compressed by the proximal aortic arch. When common arterial trunk repair was performed, the dominant aortic component of the common trunk incorporating both branch pulmonary arteries was cylindrically resected. End-to-end anastomosis of truncal root and distal ascending aorta was therefore performed to anteriorly retract the proximal arch, thereby relieving bronchial stenosis. The patient experienced an uneventful postoperative course, and postoperative computed tomography showed widely patent right main bronchus.

Patients with variants in CUL4B exhibit syndromic intellectual disability (MIM #300354). A seven-year-old boy presented with intellectual disability, a history of seizure, characteristic facial features, and short stature. Whole-exome sequencing detected a c.974+3A>G variant in CUL4B , which was subsequently confirmed to disrupt mRNA splicing. The current patient showed less pronounced phenotypic features compared with the previously reported cases. This report, therefore, provides evidence of genotype-phenotype correlations in CUL4B -related disorders., Competing Interests: Conflict of interestThe authors declare that they have no conflict of interest., (© The Author(s) 2019.)

Alveolar rhabdomyosarcoma (ARMS) is a nonepithelial tumor with skeletal muscle differentiation and typically affects adolescents and young adults. The cytological features of ARMS in body fluid have not been well characterized, which complicates diagnosis. Here, we describe the cytological features of ARMS in the ascites of a 50-year-old woman with an intra-abdominal mass and abundant ascites. Aspiration cytology of ascitic fluid revealed numerous small discohesive round cells with mild nuclear atypia and prominent nucleoli. Rhabdomyoblastic cells, characteristic of rhabdomyosarcoma, were identified rarely. Cannibalism and 'window' formation, as seen in reactive mesothelial cells, complicated the diagnosis of ARMS. Histological examination established the diagnosis of ARMS, which was confirmed by immunohistochemical expression of myogenic markers. When diagnosing ARMS from effusion samples, the diagnostic problems associated with the morphological similarity of ARMS cells to reactive mesothelial cells should be considered.

There have been increasing number of reports of SZT2-related neurological diseases, the main symptoms of which are epilepsy, developmental delay, macrocephaly and a dysmorphic corpus callosum. SZT2 functions as a regulator of mechanistic target of rapamycin complex 1 (mTORC1) signaling in cultured human cell lines and mouse tissues. However, it remains to be determined whether mutations in SZT2 in human patients alter mTORC1 signaling. In this study, we aimed to investigate the functional consequence of biallelic SZT2 variants in Epstein-Barr virus-induced lymphoblastoid cell lines (LCLs) established from two patients with a typical SZT2-related neurodevelopmental disease. Increased phosphorylation of S6 kinase and S6 was identified in patient-derived cell lines under amino acid-starved condition, suggestive of constitutive hyperactivation of mTORC1 signaling. This result was validated by constitutive lysosomal localization of mTOR in patients' LCLs. Furthermore, patients' LCLs display an excessive response to slight amino acid stimulation. Our data suggest the loss-of-function nature of SZT2 mutations in the patients, and consequent hyperactivation of mTORC1 signaling in response to both amino acid starvation and stimulation in their LCLs. By these functional analyses, the pathogenicity of newly identified SZT2 variants can be determined, allowing for more detailed characterization of genotype-phenotype correlations., Competing Interests: The authors have declared that no competing interests exist.

Background: Cerebellar ataxia, Areflexia, Pes cavus, Optic atrophy and Sensorineural hearing loss (CAPOS) syndrome is a known ATP1A3-related disorder, but little has been elucidated regarding its pathophysiology. We now report two new patients, a Japanese boy and his mother with a pathogenic mutation (c.2452G>A) in ATP1A3, who were diagnosed with CAPOS syndrome., Methods: After febrile illnesses at 7 months of age, and again at 22 months of age, the boy had a reduced level of consciousness, truncal ataxia and eye movement-disorders. The patient's 32-year-old mother may have experienced an episode of acute encephalopathy in her childhood and sustained sensorineural hearing loss. In the present study, we demonstrated chronological dynamic changes in cerebral blood flow (CBF) in the son, using serial single-photon emission computed tomography (SPECT)., Results: The serial CBF-SPECT findings using statistical methods showed progressive hyperperfusion in the frontal lobes, basal ganglia and thalamus, and hypoperfusion in the occipital and temporal lobes during the acute and subacute phases. Thereafter, the dynamic changes of CBF improved in the chronic but hypoperfusion in thalamus appeared to the chronic phase., Conclusion: The abnormal cortico-subcortical CBF may contribute to an acute encephalopathy-like condition in the acute stage of CAPOS syndrome. CAPOS syndrome is not often reported, and is possibly an under-recognized syndrome in clinically mild cases., (Copyright © 2019 The Japanese Society of Child Neurology. Published by Elsevier B.V. All rights reserved.)

Unlike most bivalve shellfishes, giant clams (tridacnines) harbor symbiotic microalgae (zooxanthellae) in their fleshy bodies. Zooxanthellae are not maternally inherited by tridacnine offspring, hence, the larvae must acquire zooxanthellae from external sources, although such algal populations or sources in the environment are currently unknown. It is well known that giant clams expel fecal pellets that contain viable zooxanthellae cells, but whether these cells are infectious or just an expelled overpopulation from the giant clams has not been investigated. In this study, we observed the ultrastructural and photosynthetic competencies of zooxanthellae in the fecal pellets of Tridacna crocea and further tested the ability of these cells to infect T. squamosa juveniles. The ultrastructure of the zooxanthellae cells showed that the cells were intact and had not undergone digestion. Additionally, these zooxanthellae cells showed a maximum quantum yield of photosystem II (Fv/Fm) as high as those retained in the mantle of the giant clam. Under the assumption that feces might provide symbionts to the larvae of other giant clams, fecal pellets from Tridacna squamosa and T. crocea were given to artificially hatched 1-day-old T. squamosa larvae. On the 9th day, 15-34% of the larvae provided with the fecal pellets took up zooxanthellae in their stomach, and on the 14th day, zooxanthellae cells reached the larval margin, indicating the establishment of symbiosis. The rate reaching this stage was highest, ca. 5.3%, in the larvae given whole (nonhomogenized) pellets from T. crocea. The composition of zooxanthellae genera contained in the larvae were similar to those in the fecal pellets, although the abundance ratios were significantly different. This study is the first to demonstrate the potential of giant clam fecal pellets as symbiont vectors to giant clam larvae. These results also demonstrate the possibility that fecal pellets are a source of zooxanthellae in coral reefs., Competing Interests: The authors have declared that no competing interests exist.

Background: Poor left ventricular (LV) growth and diastolic dysfunction long after simple total anomalous pulmonary venous connection (TAPVC) repair has been well documented and is believed to originate from insufficient preoperative volume preload. The objective of the study was to confirm these findings., Methods: Of 61 patients undergoing simple TAPVC repair between 1996 and 2016, 42 patients undergoing postoperative catheter examinations were enrolled. The mean age at the time of repair was 39 ± 117 days. Postoperative catheter examinations were conducted at a mean duration of 1.1 years after the repair. LV end-diastolic volume (LVEDV) was calculated by biplane cineangiography with the use of Simpson's method as a Graham modification., Results: LV ejection fraction, LV end-diastolic pressure (LVEDP), and cardiac index were 72% ± 5%, 10.3 ± 2.7 mm Hg, and 4.0 ± 0.7 L •·min -1 • m -2 , respectively. LVEDV was 102% ± 16% of the predictive normal value, and it strongly correlated with the predictive normal value of LV end-diastolic diameter (LVEDd) calculated by two-dimensional echocardiography (R 2  = 0.29, p = 0.005). LVEDd maintained a normal range thereafter during the entire follow-up period. LVEDP correlated linearly with age at the time of TAPVC repair (R 2  = 0.18, p = 0.007) but not with LVEDV (p = 0.67)., Conclusions: LV size maintained a normal range after the repair of simple TAPVC. High LVEDP was frequently observed a year after repair; it did not correlate with LV size but had a strong negative correlation with age at the time of repair., (Copyright © 2019 The Society of Thoracic Surgeons. Published by Elsevier Inc. All rights reserved.)

Lithium is one of the classical drugs that have been widely used for treating bipolar disorder. However, several cardiac side effects including sick sinus syndrome, bundle branch block, ventricular tachycardia/fibrillation, non-specific T-wave abnormalities in addition to Brugada-type electrocardiographic changes have been noticed in patients who were given antidepressant, anticonvulsant, and/or antipsychotic drugs besides lithium. In this study, we assessed cardiohemodynamic and electrophysiological effects of lithium carbonate by itself to begin to analyze onset mechanisms of its cardiovascular side effects. Lithium carbonate in intravenous doses of 0.1, 1, and 10 mg/kg over 10 min was cumulatively administered with an interval of 20 min to the halothane-anesthetized beagle dogs (n = 4), which provided peak plasma Li + concentrations of 0.02, 0.18, and 1.79 mEq/L, respectively, reflecting sub-therapeutic to toxic concentrations. The low and middle doses prolonged the ventricular effective refractory period at 30 min and for 5-30 min, respectively. The high dose decreased the heart rate for 45-60 min, delayed the intraventricular conduction for 15-20 min and the ventricular repolarization at 45 min, and prolonged the effective refractory period for 5-60 min. No significant change was detected in the other cardiovascular variables. Thus, lithium alone may have a wide safety margin against hemodynamic adverse events; however, it would directly and/or indirectly inhibit Na + and K + channels, which may synergistically increase the ventricular refractoriness from the sub-therapeutic concentration and decrease the heart rate at the supra-therapeutic one. These findings may partly explain its clinically observed various types of arrhythmias as well as electrocardiographic changes.

Purpose: Cancer chemotherapies have improved the prognosis of cancer patients in recent years; however, their side effects on the cardiovascular systems have emerged as a major concern in the field of both cardiology and oncology. In particular, multi-targeted tyrosine kinase inhibitors are known to induce various types of cardiovascular adverse events including hypertension, QT-interval prolongation and heart failure, but their underlying mechanisms remain elusive. To explore how to better predict such drug-induced cardiovascular adverse events, we assessed the electropharmacological effects of sunitinib using the halothane-anesthetized dogs (n = 5), while plasma concentrations of cardiac enzymes including aspartate aminotransferase, lactate dehydrogenase, creatinine kinase and cardiac troponin I  were measured., Methods: Sunitinib was intravenously administered at 0.01 and 0.1 mg/kg for 10 min with 20 min interval., Results: Sunitinib decreased the amplitude of maximum downstroke velocity of the left ventricular pressure, prolonged the isovolumic relaxation time and increased the left ventricular end-diastolic pressure in a dose-related manner without affecting the other cardiohemodynamic and electrophysiological variables. More importantly, sunitinib significantly elevated cardiac troponin I level for 30-60 min after the high dose without altering the other biomarkers., Conclusions: Monitoring of the cardiac diastolic function together with cardiac troponin I after the start of sunitinib administration may become a reliable measure to predict the onset of sunitinib-induced cardiovascular adverse events.

Introduction: Filamin A (FLNA) is located in Xq28, and encodes the actin binding protein, filamin A. A mutation in FLNA is the most common cause of periventricular nodular heterotopia (PVNH), but a clear phenotype-genotype correlation has not been established. Indeed, some patients with a FLNA mutation have recently been shown to additionally have Ehlers-Danlos-like collagenopathy or macrothrombocytopenia. In an attempt to establish a clearer correlation between clinical symptoms and genotype, we have investigated a phenotype that involves thrombocytopenia in a patient with a truncation of the FLNA gene., Case Report: We present the case of a 4-year-old girl who, at birth, showed a ventral hernia. At 2 months of age, she was diagnosed with patent ductus arteriosus (PDA) and aortic valve regurgitation. At 11 months, she underwent ligation of the PDA. She was also diagnosed with diaphragmatic eventration by a preoperative test. At 19 months, motor developmental delay was noted, and brain MRI revealed bilateral PVNH with mega cisterna magna. Presently, there is no evidence of epilepsy, intellectual disability or motor developmental delay. She has chronic, mild thrombocytopenia, and a platelet count that transiently decreases after viral infection. Dilation of the ascending aorta is progressing gradually. Genetic testing revealed a de novo nonsense heterozygous mutation in FLNA (NM&#95;001456.3: c.1621G > T; p.Glu541Ter). Immunofluorescence staining of a peripheral blood smear showed a lack of filamin A expression in 21.1% of her platelets. These filamin A-negative platelets were slightly larger than her normal platelets., Conclusion: Our data suggests immunofluorescence staining of peripheral blood smears is a convenient diagnostic approach to identify patients with a FLNA mutation, which will facilitate further investigation of the correlation between FLNA mutations and patient phenotype., (Copyright © 2018 The Japanese Society of Child Neurology. Published by Elsevier B.V. All rights reserved.)

Anti-atrial fibrillatory, proarrhythmic and cardiodepressive profiles of dronedarone were analyzed using the halothane-anesthetized beagle dogs (n = 4) to create a standard protocol for clarifying both efficacy and adverse effects of anti-atrial fibrillatory drugs. Intravenous administration of dronedarone hydrochloride in doses of 0.3 and 3 mg/kg over 30 s attained the peak plasma concentrations of 61 and 1248 ng/mL, respectively, reflecting sub- to supra-therapeutic ones. The low dose decreased the left ventricular contraction and mean blood pressure, which were enhanced at the high dose. The high dose also decreased the heart rate and cardiac output, but increased the total peripheral resistance and left ventricular end-diastolic pressure, showing its potent cardiodepressive profile. Moreover, the high dose delayed the atrioventricular nodal and intraventricular conductions in addition to the ventricular repolarization, suggesting its inhibitory action on the Ca 2+ , Na + and K + channels in the in situ heart, respectively. The high dose also prolonged the effective refractory period 1.9 times greater in the atrium than in the ventricle, explaining its clinically demonstrated efficacy against the atrial arrhythmias. Dronedarone significantly prolonged the T peak -T end in a dose-related manner with a tendency to prolong the terminal repolarization period and J-T peak c, indicating considerable risk to induce torsade de pointes. No significant change was detected in the P-wave duration by either dose, indicating the lack of effect on the atrial Na + channel in vivo. The current experimental protocol and the results of dronedarone can be used as a guide for safety pharmacological evaluation of new anti-atrial fibrillatory drugs.

Effects of moxifloxacin on QTc as well as proarrhythmic surrogate markers including J-T peak c, T peak -T end and short-term variability (STV) of repolarization were examined by using both standard E14 time-based evaluation and exposure-response modeling. The study was conducted with a single-blind, randomized, single-dose, placebo-controlled and two-period cross-over design in healthy Filipino subjects. QT interval was corrected by Fridericia's formula (QTcF). In the E14 time-based evaluation of ECG data, the largest ΔΔQTcF with 90% confidence interval was 14.1 ms (11.2-16.9) with C max of 3.39 μg/mL at 3 h post-dose (n = 69; male: 35, female: 34), indicating a positive effect on the QTcF. Moxifloxacin significantly increased the ΔΔJ-T peak c and ΔΔT peak -T end , whereas the ΔΔSTV was not altered. Meanwhile in the exposure-response modeling of the same ECG data, the slope of moxifloxacin plasma concentration-ΔΔQTcF relationship was 4.84 ms per μg/mL and the predicted ΔΔQTcF with 90% confidence interval was 13.8 ms (13.1-15.1) at C max , also indicating a positive effect on the QTcF. Importantly, results in each proarrhythmic surrogate marker obtained by the exposure-response modeling also showed high similarity to those obtained by the E14 statistical evaluation. Thus, these results of moxifloxacin may become a guide to estimate proarrhythmic potential of new chemical entities., (Copyright © 2018 The Authors. Production and hosting by Elsevier B.V. All rights reserved.)

Patients with a mutation at Arg756 in ATP1A3 have been known to exhibit a distinct phenotype, characterized by prolonged weakness and encephalopathy, triggered by febrile illness. With only eight reports published to date, more evidence is required to correlate clinical features with a mutation at Arg756. Here we report an additional case with an Arg756Cys mutation in ATP1A3. A four-year-old boy showed mild developmental delay with recurrent paroxysmal episodes of weakness and encephalopathy from nine months of age. Motor deficits, which included bilateral hypotonia, ataxia, dysmetria, limb incoordination, dysarthria, choreoathetosis, and dystonia, were observed from one year and three months. Whole-exome sequencing detected a heterozygous de novo variant at c.2266C>T (p.Arg756Cys) in ATP1A3. The episodic course and clinical features of this case were consistent with previously reported cases with mutations at Arg756. Furthermore, his phenotype of marked ataxia was more similar to that of an Arg756Cys patient with relapsing encephalopathy and cerebellar ataxia syndrome, than to those with Arg756His and Arg756Leu mutations. This report therefore provides evidence of genotype-phenotype correlations in ATP1A3-related disorders as well as in patients with mutations at Arg756 in ATP1A3., (Copyright © 2017 The Japanese Society of Child Neurology. Published by Elsevier B.V. All rights reserved.)

Mutations in SZT2 were first reported in 2013 as a cause of early-onset epileptic encephalopathy. Because only five reports have been published to date, the clinical features associated with SZT2 remain unclear. We herein report an additional patient with biallelic mutations in SZT2. The proband, a four-year-old girl, showed developmental delay and seizures from two years of age. Her seizures were not intractable and readily controlled by valproate. She showed mildly dysmorphic facies with macrocephaly, high forehead, and hypertelorism, and also had pectus carinatum. An EEG showed epileptic discharges which rarely occurred. A brain MRI revealed a short and thick corpus callosum. Whole-exome sequencing detected compound heterozygous biallelic mutations (c.8596dup (p.Tyr2866Leufs ∗ 42) and c.2930-17&#95;2930-3delinsCTCGTG) in SZT2, both of which were novel and predicted to be truncating. This case suggested a broad phenotypic spectrum arises from SZT2 mutations, forming a continuum from epileptic encephalopathy and severe developmental delay to mild intellectual disability without epilepsy. The characteristic thick and short corpus callosum observed in 7/8 cases with epilepsy, including the proband, but not in three non-syndromic cases, appears to be specific, and thus useful for indicating the possibility of SZT2 mutations. This feature has the potential to make loss of SZT2 a clinically discernible disorder despite a wide clinical spectrum., (Copyright © 2017 The Japanese Society of Child Neurology. Published by Elsevier B.V. All rights reserved.)

Microminipigs are expected as a novel animal model for cardiovascular pharmacological experiments. Since inherent vulnerability of coronary circulation of microminipigs has not been characterized, we performed dipyridamole-stress test to both microminipigs and beagle dogs, and compared the results. Dipyridamole in doses of 0.056 and 0.56 mg/kg were intravenously infused over 10 min (n = 4 for each animal). Dipyridamole decreased the systolic/diastolic blood pressures and double product in dogs as well as in microminipigs; but it did not significantly alter the heart rate or the global balance between the myocardial oxygen demand and supply in either animal. While organic coronary arterial stenosis was not detected in either animal, dogs have well-developed epicardial intracoronary networks unlike microminipigs. Like in humans, dipyridamole did not affect the ST segment of microminipigs, whereas it substantially depressed that in dogs. The results indicate the onset of subendocardial ischemia by dipyridamole in dogs may be partly associated with their well-developed native coronary collateral channels. Microminipigs would be more useful to evaluate the drugs which may affect the coronary circulation in the pre-clinical study than dogs., (Copyright © 2018 The Authors. Production and hosting by Elsevier B.V. All rights reserved.)

Amitriptyline has been reported to induce long QT syndrome in addition to Brugada syndrome. We qualitatively and quantitatively analyzed the potential of amitriptyline to induce these lethal syndromes by using the halothane-anesthetized dogs (n = 6). Amitriptyline was intravenously administered in doses of 0.1, 1 and 10 mg/kg over 10 min every 20 min, which would provide approximately 1, 10 and 100 times higher plasma concentrations than a therapeutic one, respectively. The low dose hardly altered any of the cardiovascular variables. The middle dose increased the heart rate, cardiac output and left ventricular contractility, but decreased the total peripheral vascular resistance and left ventricular end-diastolic pressure, whereas it did not alter any of the electrocardiographic variables. The high dose decreased the mean blood pressure and left ventricular contractility; suppressed atrioventricular nodal and intraventricular conduction; shortened the repolarization period without altering the J-T peak c and T peak -T end ; and prolonged the effective refractory period, providing post-repolarization refractoriness in addition to the enhancement of the middle dose-induced cardiovascular effects. Thus, amitriptyline at up to 100 times its therapeutic concentration may not be associated with the onset of long QT syndrome, but may induce Brugada syndrome.

In order to better understand the variability of pharmacodynamic and pharmacokinetic profiles of terfenadine between the previous studies as well as to qualitatively and quantitatively examine the proarrhythmic potential of its major active metabolite fexofenadine in comparison with that of terfenadine, we directly compared their electropharmacological effects with halothane-anesthetized dogs (n = 3). For this purpose, we adopted a cross-over design, which can directly compare the effects of terfenadine and fexofenadine under the identical metabolic condition. Terfenadine in doses of 0.03 and 0.3 mg/kg increased the mean blood pressure, but that of 3 mg/kg decreased it. Terfenadine also increased the heart rate and ventricular contractility in a dose-related manner; but delayed the atrioventricular nodal and intraventricular conductions as well as repolarization suggesting its proarrhythmic potential. Meanwhile, fexofenadine in the same dose increased the mean blood pressure in a dose-related manner without affecting any of the electrophysiological variables in the same animals that proarrhythmic risk of terfenadine was confirmed, indicating its lack of proarrhythmic risk. Peak plasma concentrations for fexofenadine were 3.7, 8.1 and 11.2 times greater than for terfenadine at each matching dose, indicating terfenadine may be metabolized much faster than fexofenadine. Taken together, after the low and middle doses of terfenadine, vasopressor effect of a metabolite fexofenadine could be greater than the depressor effect of parent compound terfenadine, but its reverse would be correct after the high dose. Thus, the cross-over analysis can be an effective way to better understand drug-induced cardiovascular responses.

A beverage made of red wine vinegar and grape juice (Yamanashi-no-megumi™) was developed as a supplemental fluid containing polyphenols, which has been clinically shown to enhance the colonic transit. In this study, we assessed the mechanism of its prokinetic action by analyzing the effects on both the colonic phosphodiesterase activity of rats (n=4) and the isolated colonic strip preparation of guinea pigs (n=4). The 7% (v/v) solution of the beverage significantly decreased the phosphodiesterase activity by 9% (n=4). The beverage in concentrations of 0.7, 2.1 and 7% (v/v) relaxed the colonic strips pre-contracted by 1 µmol/L of carbachol in a concentration-related manner with 50, 58 and 79%, each response of which was diminished to 11, 19 and 46%, respectively in the presence of 100 µmol/L of L-nitro-arginine methyl ester. These results obtained by biochemical, functional and pharmacological analyses suggest that the beverage could relax the colon through both cAMP-associated and nitric oxide-dependent pathways, which may partly explain clinically observed prokinetic effect of the beverage.

We analyzed electropharmacological characteristics of microminipigs under halothane-anesthesia using anti-influenza virus drug oseltamivir, which has been known to possess multi-channel blocking properties, including Na + , Ca 2+ and K + channels (n = 4). Oseltamivir in doses of 0.3, 3 and 30 mg/kg was intravenously infused over 10 min with an interval of 20 min, which provided peak plasma concentrations 1.4, 7.4 and 125.5 µg/mL, respectively. The low dose did not alter any of the cardiovascular variables. The middle dose decreased the heart rate at 30 min after the start of the infusion. The high dose transiently returned the heart rate toward the baseline for 10-15 min, but decreased it for 20-60 min; decreased the mean blood pressure for 5-60 min; prolonged the PR interval for 10-60 min, and the QRS width for 10-20 min; but shortened the QT interval for 10-30 min, and the QTc for 5-60 min. Thus, oseltamivir can suppress the sinus automaticity, and atrioventricular nodal and intraventricular conduction; and decrease the mean blood pressure, extents of which were greater in microminipigs than in beagle dogs in our previous observation in spite of similar plasma concentrations, reflecting higher sensitivity of microminipigs for Na + and Ca 2+ channel inhibition than that of beagle dogs. In contrast to beagle dogs, oseltamivir shortened the repolarization period in microminipigs, indicating that oseltamivir can more potently inhibit the inward currents than the outward ones in the hearts of microminipigs. This information may help improve utilizatione of microminipigs as a laboratory animal.

Background: The influence of S-carboxymethylcystein (S-CMC) on the proliferation ability of goblet cells in nasal polyp epithelium in response to inflammatory stimulation was examined., Methods: The subjects were patients with chronic paranasal sinusitis. An epithelial cell culture system was established using nasal polyp mucosa excised during endoscopic paranasal sinus surgery. The samples were divided into 4 groups (group a: control group, group b: 10 ng/mL tumor necrosis factor-α (TNF-α) treatment group, group c: 10 -7 M S-CMC and 10 ng/mL TNF-α treatment group, group d: 10 -5 M S-CMC and 10 ng/mL TNF-α treatment group). The total number of epithelial cells and number of goblet cells were measured under a microscope, and the ratio of goblet cells to the total number of epithelial cells was calculated., Results: In group b, 10 ng/mL of TNF-α significantly increased the number of goblet cells compared with group a, suggesting involvement of TNF-α in goblet cell proliferation. In addition, the number of goblet cells significantly decreased in group d compared with that in group b, and it also decreased in group c compared with that in group b, although the difference was not significant, and the decrease was smaller than that in group d, suggesting that S-CMC inhibited goblet cell proliferation in a concentration-dependent manner., Conclusion: TNF-α promoted goblet cell proliferation in nasal polyps, suggesting its influence on nasal polyp formation. As S-CMC inhibited inflammatory stimulation-induced goblet cell proliferation in nasal polyp epithelium, it may be useful for the treatment of sinusitis.

Using moxifloxacin and terfenadine, which are known to induce benign and malignant QT interval prolongation, respectively, we analysed whether halothane-anaesthetized microminipigs are an appropriate model for assessing the risk of drug-induced long QT syndrome. Moxifloxacin (0.03, 0.3 and 3 mg/kg) and terfenadine (0.03, 0.3 and 3 mg/kg) were intravenously infused over 10 min. with a pause of 20 min. to the halothane-anaesthetized microminipigs (n = 4 for each drug). Moxifloxacin decreased the heart rate, whereas it increased the blood pressure in a dose-related manner. It also prolonged the PR interval and QT/QTc in a dose-related manner without altering the QRS width. Terfenadine decreased the heart rate and blood pressure, whereas it prolonged the PR interval, QRS width and QT/QTc in a dose-related manner. Terfenadine significantly prolonged the beat-to-beat variability of QT interval reflecting its pro-arrhythmic potential, which was not observed with moxifloxacin. The peak plasma concentrations of moxifloxacin and terfenadine after doses of 3 mg/kg were 4.81 and 10.15 μg/mL, respectively, which were both 1.5 times less in microminipigs than those previously reported in dogs. These results indicate that halothane-anaesthetized microminipigs would be useful for detecting drug-induced cardiovascular responses as well as differentiating benign from malignant QT interval prolongation like dogs, although there may be some differences in pharmacokinetic profile between these animals., (© 2017 Nordic Association for the Publication of BCPT (former Nordic Pharmacological Society).)

Novel thienopyrimidine compounds 2 and 3 were discovered from high-throughput screening as Natriuretic Peptide Receptor A (NPR-A) agonists. Scaffold hopping of a thienopyrimidine ring to a quinazoline ring, introduction of the basic functional group and optimization of the substituent on the 6-position of the benzene ring of quinazoline led to improved agonistic activity. We discovered compound 48, which showed potent agonistic activity for NPR-A with an EC 50 value of 0.073μM, indicating 350-fold potency compared to the hit compound 3., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) have been used in many studies to assess proarrhythmic risks of chemical compounds. In those studies, field potential durations (FPD) of hiPSC-CMs have been corrected by clinically used Fridericia's and/or Bazett's formulae, however, the rationale for the use of these formulae has not been well established. In the present study, we developed a correction formula for experiments using hiPSC-CMs. First, we analyzed the effect of beating rate on FPD in the hiPSC-CMs sheets with electrical stimuli and a HCN channel inhibitor zatebradine. Next, we examined the relationship between the electrophysiological properties and the expression levels of ion channel genes in the cell sheets. Zatebradine slowed the beating rate and allowed to analyze FPD changes at various pacing cycle lengths. Rate-dependent change in the repolarization period was smaller in the cell sheets than that reported on the human hearts, which can be partly explained by lower gene expression level of hKCNJ2 and hKCNE1. Thus, non-linear equation for correcting FPD in the cell sheet; FPDc = FPD/RR 0.22 with RR given in second was obtained, which may make it feasible to assess net repolarization delay by various chemical compounds with a chronotropic action., (Copyright © 2017 The Authors. Production and hosting by Elsevier B.V. All rights reserved.)

Fatal cases with the use of atypical antipsychotic drug paliperidone have been reported; however, there was no clinical report describing paliperidone-induced torsade de pointes. In this study we assessed its electropharmacological effects together with its proarrhythmic potential in intravenous doses of 0.03, 0.3 and 3 mg/kg using the halothane-anesthetized dogs (n = 5), which could provide approximately 2, 20 and 200 times higher peak plasma drug concentrations than its therapeutic level, respectively. Paliperidone exerted potent vasodilator effect resulting in hypotension, which may be largely explained by its α 1 -adrenoceptor blocking action. In vivo electrophysiological results suggest that paliperidone may inhibit human ether-à-go-go-related gene K + channel in a dose-related manner and modestly suppress Na + channel in the in situ heart. The high dose of paliperidone may have some potential to induce early afterdepolarization that can trigger lethal ventricular arrhythmias, whereas the low and middle doses lack such proarrhythmic possibility, indicating that at least 20 times higher plasma concentration may be considered to be safe., (Copyright © 2017 The Authors. Production and hosting by Elsevier B.V. All rights reserved.)

Background: Potential cardiovascular benefits of precordial percussion pacing (PPP) during cardiac standstill are unknown.Methods and Results:A cardiac standstill model in amicrominipigwas created by inducing complete atrioventricular block with a catheter ablation technique (n=7). Next, the efficacy of cardiopulmonary resuscitation by standard chest compressions (S-CPR), PPP and ventricular electrical pacing in this model were analyzed in series (n=4). To assess the mechanism of PPP, a non-selective, stretch-activated channel blocker, amiloride, was administered during PPP (n=3). Peak systolic and diastolic arterial pressures during S-CPR, PPP and ventricular electrical pacing were statistically similar. However, the duration of developed arterial pressure with PPP was comparable to that with ventricular electrical pacing, and significantly greater than that with S-CPR. Amiloride decreased the induction rate of ventricular electrical activity by PPP in a dose-related manner. Each animal survived without any neurological deficit at 24, 48 h and 1 week, even with up to 2 h of continuous PPP., Conclusions: In amicrominipigmodel of cardiac standstill, PPP can become a novel means to significantly improve physiological outcomes after cardiac standstill or symptomatic bradyarrhythmias in the absence of cardiac pacing. Activation of the non-selective stretch-activated channels may mediate some of the mechanophysiological effects of PPP. Further study of PPP by itself and together with S-CPR is warranted using cardiac arrest models of atrioventricular block and asystole.

Since an antipsychotic drug haloperidol has been clinically reported to induce QT interval prolongation and torsade de pointes, in this study its risk stratification for the onset of torsade de pointes was performed by using the chronic atrioventricular block canine model with a Holter electrocardiogram. Haloperidol in a dose of 3 mg kg -1 p.o. prolonged the QT interval, but it did not induce torsade de pointes during the observation period of 21 h (n = 4), indicating that the dose would be safe. Meanwhile, haloperidol in a dose of 30 mg kg -1 p.o. significantly increased the short-term variability in beat-to-beat analysis of QT interval (n = 4), and it induced torsade de pointes in 4 animals out of 4, showing that the dose could be torsadogenic. Since 3 mg kg -1 p.o. of haloperidol in this study can be estimated to provide about 8 times higher plasma concentrations than its therapeutic level, haloperidol may be used safely for most of the patients, as long as its plasma drug concentration is kept within the therapeutic range.

Anti-atrial fibrillatory and proarrhythmic potentials of amiodarone were simultaneously analyzed by using the halothane-anesthetized beagle dogs (n = 4) in order to begin to prepare standard protocol for clarifying both efficacy and adverse effects of anti-atrial fibrillatory drugs. Intravenous administration of 0.3 mg/kg of amiodarone hydrochloride decreased the heart rate and mean blood pressure. Additional administration of 3 mg/kg of amiodarone hydrochloride prolonged the QT interval besides the effects observed by the low dose, whereas it showed 1.6 times larger prolongation in the effective refractory period of the atrium than that of the ventricle, which may explain its clinical efficacy against atrial arrhythmias. However, no significant change was detected by either dose in the early repolarization assessed by corrected J-T peak or the late repolarization done by T peak -T end in the electrocardiogram, although the former tended to be shortened and the reverse was true for the latter. Lack of prolongation in the early repolarization will make it feasible to better understand why amiodarone lacks proarrhythmic potential in spite of the QT-interval prolongation. Thus, these results of amiodarone obtained by current protocol may become a guidance on assessing efficacy and adverse effects of new anti-atrial fibrillatory drugs in vivo.

We pharmacologically characterized microminipigs as an in vivo experimental model by assessing cardiovascular effects of pilsicainide, verapamil and E-4031, which can preferentially inhibit cardiac Na + , Ca 2+ and K + channels, respectively. Intravenous infusion of 1 mg/kg of pilsicainide (n = 4), 0.1 mg/kg of verapamil (n = 4) and 0.01 followed by 0.1 mg/kg of E-4031 (n = 5) over 10 min decreased the heart rate, mean blood pressure and ventricular contractility. Moreover, pilsicainide prolonged the PR interval, QRS width and QTc; verapamil prolonged the PR interval, but shortened the QRS width and QTc; and E-4031 prolonged the QTc, whereas no substantial change was detected in the PR interval or QRS width. Peak plasma concentrations of pilsicainide, verapamil and E-4031 in microminipigs were 1.7-4.8 times higher than those expected in humans and dogs, possibly due to smaller effective volume of drug distribution. The extent of the drug-induced cardiovascular responses was generally greater in microminipigs than in humans and dogs, which could be explained by the following possibilities; namely unique pharmacokinetic profile, less great reflex-mediated increase of sympathetic tone and/or smaller repolarization reserve in microminipigs. These information may make it feasible to apply this new-type animal to a tool for assessing cardiac safety profiles of new chemical entities., (Copyright © 2017 The Authors. Production and hosting by Elsevier B.V. All rights reserved.)

A 53-year-old woman with end-stage renal disease and hypertension, who had received regular artificial hemodialysis for 10 years, has been treated with candesartan in a dose of 8 mg/day against her hypertension, but premature ventricular contractions were often observed during the hemodialysis. QT interval was 445 ms before hemodialysis, which was prolonged to 515 ms immediately after it, possibly reflecting the presence of reduced repolarization reserve in her heart. Since the blood pressure was often elevated to >160 mmHg before the hemodialysis, a daily dose of an L/N-type Ca 2+ channels blocker cilnidipine of 5 mg/day was added. Three months later, the electrocardiogram was obtained before hemodialysis, revealing the basal QT interval was shortened to 416 ms. More importantly, in the electrocardiogram recorded immediately after the hemodialysis, the QT interval was 429 ms, indicating that 3 months administration of cilnidipine may restore the reduced repolarization reserve. As well, we observed that premature ventricular contractions during the hemodialysis had disappeared. Thus, cilnidipine may become a new upstream therapy to reduce the risk of lethal arrhythmias.

We tried to establish the halothane-anesthetized microminipigs as an alternative animal model for non-clinical toxicity and/or safety pharmacology studies. In order to characterize the halothane-anesthetized microminipigs, we firstly clarified the effects of halothane anesthesia on their cardiovascular system (n = 5). Then, we examined the cardiovascular effects of dl-sotalol in doses of 0.1, 0.3 and 1 mg/kg, i.v. on the halothane-anesthetized microminipigs (n = 6). Induction of the halothane anesthesia by itself prolonged the QT interval as well as QTcF, suggesting that the halothane anesthesia can reduce the cardiac repolarization reserve in microminipigs like in dogs. dl-Sotalol showed more potent negative chronotropic, dromotropic and hypotensive effects together with repolarization delay in microminipigs than in dogs, although each cardiovascular response to dl-sotalol was directionally similar between them, suggesting greater basal sympathetic tone and/or smaller volume of distribution of the drug in microminipigs than in dogs. Analyses of proarrhythmic surrogate markers indicate that T peak -T end and short-term variability of QT interval may be more sensitive to detect the dl-sotalol-induced direct electrophysiological changes in microminipigs than in dogs, but its reverse will be true for J-T peak c. Thus, these results may help better understand the drug-induced cardiovascular responses in microminipigs.

In order to characterize human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) sheets as a model for detecting drug-induced conduction disturbance, we examined their electrophysiological and electropharmacological properties by using the multi-electrode array system with a programmed electrical stimulation protocol. At pre-drug control, the conduction speed, effective refractory period and field potential duration were 0.14 ± 0.01 m/sec, 453 ± 10 msec and 361 ± 9 msec, respectively at a cycle length of 1,000 msec (n = 18). Shortening the pacing cycle length from 1,000 to 600 msec decreased the conduction speed and field potential duration, but prolonged the effective refractory period. Disopyramide, lidocaine and flecainide decreased the conduction speed but prolonged the effective refractory period and field potential duration, whereas the reverse was true for verapamil. Thus, conduction properties of the cell sheet may largely depend on the extent of Na + channel availability as is the case in the human ventricle. Importantly, there was no relationship between the conduction delay and 1 st spike amplitude reduction after the treatment of Na + channel blockers. These findings may provide crucial guide on future application of this new technology for early phase safety pharmacological screening of new chemical entities.

We assessed the anti-anginal effects of cilnidipine in comparison with those of nicardipine and nifedipine (1 and 10 µg/kg, n = 6 for each drug) or vehicle (n = 6) by using the vasopressin-induced angina model of rats. The administration of vasopressin (0.5 IU/kg, i.v.) to the rats depressed the S-wave level of the electrocardiogram reflecting the presence of subendocardial ischemia, whereas it significantly increased the mean blood pressure, resulting in the decrease of the heart rate and the prolongation of the PR interval possibly through a reflex-mediated increase in vagal tone. Cilnidipine suppressed the vasopressin-induced depression of the S-wave level in a dose-related manner, which was not observed by nicardipine or nifedipine. In addition, the low dose of cilnidipine hardly affected the vasopressin-induced pressor response, but it attenuated the negative dromotropic effect, suggesting N-type Ca 2+ channel inhibition by cilnidipine might have suppressed the parasympathetic nerve activity in vivo like those reported in the sympathetic nerve. Thus, cilnidipine may become a useful strategy for inhibiting coronary vasospasm-induced anginal attack.

Direct vasodilator effects of nitroglycerin, nifedipine, cilnidipine and diltiazem on human skeletonized internal mammary artery graft harvested with ultrasonic scalpel were assessed in the presence of 0.1 or 0.2 µM of noradrenaline. Ring preparations were made of distal end section of the bypass grafts, and those dilated by acetylcholine were used for assessment. Each drug dilated the artery in a concentration-related manner (0.01-10 µM, n = 6 for each drug) with a potency of nitroglycerin > nifedipine = cilnidipine > diltiazem. These results indicate that nitroglycerin can be useful for treating internal mammary artery spasm, that clinical utility of diltiazem may not depend on its vasodilator effect on the bypass graft, and that cilnidipine as well as nifedipine will have anti-spastic action which is in the middle between those of nitroglycerine and diltiazem.